Human GRB2 knockout HCT116 cell lysate (ab275248)
Overview
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Product name
Human GRB2 knockout HCT116 cell lysate -
Product overview
Knockout cell lysate achieved by CRISPR/Cas9. -
Parental Cell Line
HCT116 -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in allele 1 and 2 bp deletion in allele 2 in exon 2 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
Reconstitution notes
To use as WB control, resuspend the lyophilizate in 50 µL of LDS* Sample Buffer to have a final concentration of 2 mg/ml. For reducing conditions, we recommend a final concentration of 0.1 M DTT.*Usage of SDS sample buffer is not recommended with these lyophilized lysates.
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Notes
Lysate preparation: Our lysates are made using RIPA buffer to which we add a protease inhibitor cocktail and phosphatase inhibitor cocktail (ratio: 300:100:10). This means that the protein of interest is denatured. If you require a native form of the protein please use the live cell version - found here. Please refer to our lysis protocol for further details on how our lysates are prepared.
User storage instructions: Lyophilizate may be stored at 4°C. After reconstitution, store at -20°C for short-term storage or -80°C for long-term storage.
Access thousands of knockout cell lysates, generated from commonly used cancer cell lines.
See here for more information on knockout cell lysates.Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.
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Tested applications
Suitable for: WBmore details
Properties
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Storage instructions
Store at -80°C. Please refer to protocols. -
Components 1 kit ab277292 - Human GRB2 knockout HCT116 cell lysate 1 x 100µg ab277289 - Human wild-type HCT116 cell lysate 1 x 100µg -
Research areas
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Cell type
epithelial -
Disease
Carcinoma -
Gender
Male
Target
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Function
Adapter protein that provides a critical link between cell surface growth factor receptors and the Ras signaling pathway.
Isoform GRB3-3 does not bind to phosphorylated epidermal growth factor receptor (EGFR) but inhibits EGF-induced transactivation of a RAS-responsive element. Isoform GRB3-3 acts as a dominant negative protein over GRB2 and by suppressing proliferative signals, may trigger active programmed cell death. -
Sequence similarities
Belongs to the GRB2/sem-5/DRK family.
Contains 1 SH2 domain.
Contains 2 SH3 domains. -
Domain
The SH3 domains mediate interaction with RALGPS1 and SHB. -
Cellular localization
Golgi apparatus. - Information by UniProt
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Alternative names
- Abundant SRC homology
- Adapter protein GRB2
- ASH
see all
Associated products
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KO cell lines
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab275248 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| WB |
Use at an assay dependent concentration. Predicted molecular weight: 25 kDa.
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| Notes |
|---|
|
WB
Use at an assay dependent concentration. Predicted molecular weight: 25 kDa. |
Images
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Western blot - Human GRB2 knockout HCT116 cell lysate (ab275248)
Lane 1: Wild type HCT116 (human colorectal carcinoma epithelial cell), whole cell lysate, 20 ug
Lane 2: GRB2 knockout HCT116 (ab273715) whole cell lysate, 20 ug
Lane 3: HeLa (human epithelial cell line from cervix adenocarcinoma), whole cell lysate, 20 ug
Lane 4: 293T ( human embryonic kidney epithelial cell), whole cell lysate, 20 ug
Lanes 1-4: Merged signal (red and green). Green - ab281846 observed at 26KDa. Red - loading control ab181602 (Rabbit monoclonal [EPR16891] to GAPDH) observed at 36 kDa.
Lanes 1-2: ab281846 Anti-GRB2 antibody was shown to react with GRB2 in HCT116 cells in Western blot. Loss of signal was observed when GRB2 knockout cell line ab273715 (knockout cell lysate ab275248) was used. Wild-type and GRB2 knockout samples were subjected to SDS-PAGE.
ab281846 and Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) were incubated at 4? overnight at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 680CW) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800RD) preadsorbed (ab216772) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.Blocking and diluting buffer and concentration: Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS
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Western blot - Human GRB2 knockout HCT116 cell lysate (ab275248)Lane 1: Wild-type HCT116 cell lysate 20 ug
Lane 2: GRB2 knockout HCT116 cell lysate 20 ug
Lane 3: HeLa cell lysate 20 ug
Lane 4: HEK293 cell lysate 20 ug
Lanes 1 - 4: Merged signal (red and green). Green - ab32111 observed at 25 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.
ab32111 was shown to react with GRB2 in wild-type HCT 116 cells in western blot with loss of signal observed in GRB2 knockout cell line ab273715 (GRB2 knockout cell lysate ab275248). HCT 116 wild-type and GRB2 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab32111 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging. -
Sanger Sequencing - Human GRB2 knockout HCT116 cell lysate (ab275248)Allele-1: 2 bp deletion in exon 2 . -
Sanger Sequencing - Human GRB2 knockout HCT116 cell lysate (ab275248)Allele-2: 1 bp deletion in exon 2 .
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab275248 has not yet been referenced specifically in any publications.